State of actin in gastric parietal cells.
Remodeling of the apical membrane-cytoskeleton has been suggested to occur when gastric parietal cells are stimulated to secrete HCl. The present experiments assayed the relative amounts of F-actin and G-actin in gastric glands and parietal cells, as well as the changes in the state of actin on stimulation. Glands and cells were treated with a Nonidet P-40 extraction buffer for separation into detergent-soluble (supernatant) and detergent-insoluble (pellet) pools. Two actin assays were used to quantitate actin the deoxyribonuclease I binding assay to measure G-actin and F-actin content in the two pools and a simple Western blot assay to quantitate the relative amount of actin in the pools. Functional secretory responsiveness was assayed by aminopyrine accumulation. About 5% of the total parietal cell protein is actin, with about 90% of the actin present as F-actin. Stimulation of acid secretion resulted in no measurable change in the relative amounts of G-actin and cytoskeletal F-actin. Treatment of gastric glands with cytochalasin D inhibited acid secretion and resulted in a decrease in F-actin and an increase in G-actin. No inhibition of parietal cell secretion was observed when phalloidin was used to stabilize actin filaments. These data are consistent with the hypothesis that microfilamentous actin is essential for membrane recruitment underlying parietal cell secretion. Although the experiments do not eliminate the importance of rapid exchange between G- and F-actin for the secretory process, the parietal cell maintains actin in a highly polymerized state, and no measurable changes in the steady-state ratio of G-actin to F-actin are associated with stimulation to secrete acid.
Forte
J G
JG
Department of Molecular and Cell Biology, University of California, Berkeley 94720, USA.
Ly
B
B
Rong
Q
Q
Ogihara
S
S
Ramilo
M
M
Agnew
B
B
Yao
X
X
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002463:Cell Membrane Permeability; D002927:Cimetidine / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000502:physiology; D005704:Gallbladder / Q000166:cytology / Q000502:physiology; D005753:Gastric Mucosa / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D006632:Histamine / Q000494:pharmacology; D066298:In Vitro Techniques; D007700:Kinetics; D007962:Leukocytes / Q000502:physiology; D010295:Parietal Cells, Gastric / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D011817:Rabbits
D000199:Actins; D002927:Cimetidine; D006632:Histamine